Abstract

A concept that has arisen over the last decade is that proteins can, in general, be covalently modified by polypeptides, resulting
in alterations in their fate and function. The first‐identified and most well studied of these modifying polypeptides is ubiquitin.
Although targeting for proteasomal degradation is the best studied outcome of ubiquitylation, we now understand that modification
of proteins with ubiquitin has numerous other cellular roles that alter protein function and that are unrelated to proteasomal
degradation. Ubiquitylation is a complex process that is regulated at the level of both addition and removal of ubiquitin
from target proteins. This unit includes a number of different basic protocols that will facilitate the study of components
of the ubiquitin system and substrate ubiquitylation both in vitro and in cells. Because another protein modifier, NEDD8,
itself regulates aspects of the ubiquitin system, basic protocols on neddylation are also included in this unit.

Huibregtse, J.M.,
Scheffner, M., and
Howley, P.M.
1993.
Cloning and expression of the cDNA for E6‐AP, a protein that mediates the interaction of the human papillomavirus E6 oncoprotein with p53.
Mol. Cell Biol.
13:775‐784.